Ethyl-substituted erythromycin derivatives produced by directed metabolic engineering

Area of Science:

• Biotechnology
• Synthetic Biology
• Microbial Engineering

Background:

• Erythromycin A (ErA) is a crucial macrolide antibiotic produced by Saccharopolysporia erythraea.
• The C-6 position of the ErA polyketide backbone typically features a methyl side chain.
• Modifying this side chain could lead to novel antibiotic derivatives with altered properties.

Purpose of the Study:

• To produce a novel erythromycin derivative, 6-desmethyl-6-ethylerythromycin A (6-ethylErA), through directed genetic manipulation.
• To investigate the feasibility of replacing the C-6 methyl group with an ethyl moiety in erythromycin A.
• To enhance the production of 6-ethylErA by addressing substrate limitations in the engineered strain.

Main Methods:

• Directed genetic engineering of Saccharopolyspora erythraea.
• Replacement of the methylmalonate-specific acyltransferase (AT) domain in erythromycin polyketide synthase with an ethylmalonate-specific AT domain from niddamycin.
• Supplementation with ethylmalonate precursors.
• Expression of crotonyl-CoA reductase to enhance substrate availability.

Main Results:

• The initial genetic modification resulted in the production of ErA, not the desired 6-ethylErA.
• Supplementation with ethylmalonate precursors led to the production of small quantities of 6-ethylErA alongside ErA.
• Co-expression of crotonyl-CoA reductase significantly increased the yield, making 6-ethylErA the primary macrolide produced.

Conclusions:

• Directed genetic engineering can be used to create novel macrolide structures like 6-ethylErA.
• Substrate availability is a critical factor in the efficient production of engineered polyketides.
• Metabolic engineering strategies, including the expression of specific enzymes, can overcome production bottlenecks for novel compounds.